This invention broadly relates to devices for sensing and determining the concentration of oxygen or an oxygen-related analyte in a medium. More specifically, this invention relates to sensing elements for sensing and sensor systems for determining blood or tissue oxygen concentrations.
Oftentimes during surgical procedures, a number of blood analytes are monitored in real time. For example, during open-heart surgery, the surgeon and other members of the surgical team often monitor blood pH, as well as the concentration of various blood gases, such as O2 and CO2. It is also of interest to monitor these analytes in patients for extended periods of time before or after surgery. Furthermore, it is oftentimes desirable to monitor these analytes in critically ill patients in an intensive care unit. It may also be desirable to monitor other blood analytes, such as glucose, in critically ill patients.
Because of their unique properties, fluorescence-based sensing elements have been employed in sensor systems designed for real time monitoring of blood analytes including pH, CO2O2 and K+. The sensing element comprises a sensor film and a substrate for holding the sensor film and bringing it into contact with the patient""s blood. Typically, the sensor film comprises a fluorescent substance that is distributed in a polymeric matrix that is permeable to the analyte of interest (e.g. O2 and CO2 sensors). Alternatively, the fluorescent substance is anchored to a polymeric film that is contacted with the analyte of interest (e.g. pH and K+ sensors).
For in vivo applications, the sensor film may be disposed on the tip of an optical fiber and then inserted into an arterial catheter or into a needle for insertion into the tissue of the patient, as disclosed in Lubbers et al U.S. Pat. No. Re 31,879, and Maxwell U.S. Pat. No. 4,830,013. For ex vivo and extracorporeal applications, the sensor film may be disposed on a carrier disk and incorporated into a disposable flow through cassette, which is then placed in an arterial line circuit or an extracorporeal blood loop as shown in Cooper U.S. Pat. No. 4,640,820. Each of these patents is incorporated by reference in its entirety herein.
When exposed to light at a proper wavelength, the fluorescent substances (referred to hereinafter as xe2x80x9cfluorophoresxe2x80x9d) absorb energy and are driven from their ground state energy level into an excited state energy level. Fluorophores are unstable in their excited states and fluoresce (radiative decay) or give off thermal energy (non-radiative decay) as they return to their ground state. The fluorescence intensity, I, represents the intensity of the emission given off by the fluorophore as it returns to the ground state. The fluorescence lifetime, xcfx84, represents the average amount of time the fluorophore remains in its excited state prior to returning to the ground state.
Fluorescence based oxygen sensing elements work on the principle that oxygen molecules can collisionally quench the excited state of a fluorophore. When the fluorophore is excited in the presence of oxygen molecules, collisional interactions between the excited state and the oxygen molecule introduce a new mechanism for non-radiative decay, resulting in a decrease in both the fluorescence intensity and the excited state lifetime. Thus, blood gas monitoring systems which employ fluorescence based oxygen sensing elements have been designed to monitor oxygen-related changes in fluorescence intensity or excited state lifetime of the fluorophore.
The relationship between the fluorescence intensities and lifetimes in the absence (Io, xcfx84o) and presence (I, xcfx84) of oxygen is described by the Stern-Volmer equation:                                                                                           I                  0                                I                            =                                                τ                  0                                τ                                                                                        =                                                                    k                    q                                    ⁡                                      [                                          O                      2                                        ]                                                                                        k                    em                                    +                                      k                    nro                                                                                                                          =                              1                +                                                      k                    q                                    ⁢                                                            τ                      0                                        ⁡                                          [                                              O                        2                                            ]                                                                                                                                              =                              1                +                                                      ak                    q                                    ⁢                                      τ                    0                                    ⁢                  p                  ⁢                                      xe2x80x83                                    ⁢                                      O                    2                                                                                                                          =                              1                +                                                      K                    SV                                    ⁢                  p                  ⁢                                      xe2x80x83                                    ⁢                                      O                    2                                                                                                          Equation        ⁢                  xe2x80x83                ⁢        1            
where [O2] is the concentration of oxygen in the sensing element; pO2 is the partial pressure of oxygen in the medium being sensed; a is the solubility constant for oxygen in the sensing element which equals [O2]/pO2; kq is the bimolecular quenching constant in the sensing element; kem represents the rate constant for radiative decay; knro represents the rate constant for non-radiative decay in the absence of oxygen; and KSV is the Stern-Volmer quenching constant.
Relative fluorescence intensities (Io/I) or relative fluorescence lifetimes xcfx84o/xcfx84 are measured experimentally. Ideally, a plot of Io/I or xcfx84o/xcfx84 against pO2 should give a straight line with a slope of KSV=akqxcfx84o and an intercept of unity. A calibration curve can be made of intensity versus concentration, and from this the concentration of the quenching species in the medium can be determined.
When a disposable flow-through cassette containing a sensor disk is clipped into the optics head of a blood gas-monitoring device, there are several factors that can lead to variability in the intensity of the fluorescent return signal that is given off by the sensing element and detected by the sensor system detector. Similarly, when a fiber optic probe having a sensor film at the distal end of the fiber is inserted into an arterial catheter or into tissue, there are several factors that can lead to variability in the intensity of the fluorescent return signal. In both configurations, these sources of variability include optical coupling efficiencies throughout the optical train, optical coupling to the cassette or fiber optic probe, lamp intensity, concentration of the fluorophore in the sensing element, and thickness of the sensing element. Even after the sensor system has been calibrated, return signal intensities can drift as a result of fiber bending, fluctuations in lamp intensity, temperature dependent changes in optical coupling efficiencies or the detection electronics, and photo-bleaching of the fluorophore. The effects of fiber bending and photo-bleaching are particularly pronounced in fiber optic probes.
A well recognized advantage of using fluorescence lifetime to determine oxygen concentration is that fluorescence lifetime is insensitive to variations in sensor film thickness, optical coupling efficiencies, fiber bending, and fluctuations in lamp intensity. The two most common techniques for measuring fluorescence lifetimes are the pulse method and the phase modulation method. In the pulse method, the fluorophore is excited by a brief pulse of light, and the decay of fluorescence is determined. In the phase modulation method, the fluorophore is excited by a light beam that is preferably sinusoidally amplitude modulated at a radial frequency xcfx89=2xcfx80f, where f is the frequency in cycles per second. The fluorescence emission from the fluorophore is a forced response to this excitation signal, and is therefore amplitude modulated at the same radial frequency xcfx89 as the excitation signal. However, because of the finite lifetime of the fluorophore in the excited state, the emission is phase shifted by an angle xcex8 with respect to the excitation signal. Furthermore, the amplitude or intensity of the emission is less modulated (demodulated) by an amount m with respect to the excitation signal. The lifetime of the fluorophore can be calculated in a known manner from measurements of the phase shift (tan xcex8=xcfx89xcfx84) and the demodulation factor (m=(1+xcfx892xcfx842)xe2x88x921/2).
By measuring the phase shift, one can determine the fluorescence lifetime and therefore the analyte concentration. The Stern-Volmer slope is determined by measuring the phase shift and plotting the equation                                           τ            0                    τ                =                                            tan              ⁢                              xe2x80x83                            ⁢                              θ                0                                                    tan              ⁢                              xe2x80x83                            ⁢              θ                                =                      1            +                                          K                SV                            ⁢              p              ⁢                              xe2x80x83                            ⁢                              O                2                                                                        Equation        ⁢                  xe2x80x83                ⁢        2            
This approach still requires measurement of a reference signal from the light source or from the driver electronics, and this reference signal must be used to correct for phase drift in the detection electronics. However, it is not necessary to send the reference signal through the sensing element since the measured phase shift is independent of optical coupling losses, fiber bending, variations in dye concentration or changes in light source amplitude.
Regardless of the method used to determine the lifetimes or intensity, the slope of the resulting Stern-Volmer calibration plots will necessarily depend on the value of xcfx84o. Measured values for xcfx84o generally vary from sensing element to sensing element as a result of self-quenching and microheterogeneities of the fluorophore in the sensor films. Therefore, each sensing element must be individually calibrated using a two-point calibration method.
A procedure outlined in Bentsen U.S. Pat. No. 5,403,746 successfully addresses the two-point calibration issues for a flow-through cassette comprising intensity based sensing elements for pH, CO2 and O2. This configuration and procedure is commonly employed for extracorporeal blood gas sensing systems used during open heart surgery. This procedure is lengthy (30 min) and involves exposing the cassette to a buffer solution that is alternately exposed to two different calibration gas mixtures having different partial pressures of oxygen and carbon dioxide. The two calibrants will typically have known analyte concentrations, one close to the maximum, and the other close to the minimum concentrations of the range over which measurements are to be taken. By alternately exposing the sensing element to the two calibrants, the slope and intercept of a calibration plot may be determined so that the sensor system can accurately measure unknown concentration of blood analytes. Two point calibration involves adjusting the slope and intercept of the calibration data, as represented by the lookup table data or mathematical equation stored in memory of the sensor processor, until the relationship characterized by the data extends through the points corresponding to those of the known calibrants. A similar procedure can be applied to the calibration of single fiber sensing elements incorporated within a protective needle or inserted into an arterial catheter as taught in Maxwell U.S. Pat. No. 4,830,013.
In bedside applications, it is desirable to monitor blood gases consistently over an extended period of time. For example, it is desired to leave the sensing element in the a-line circuit for up to 72 hrs, the standard in-dwell time for an arterial catheter. Unfortunately, current sensor systems drift substantially over this period of time and require recalibration. Since two point calibration procedures require the sensing element to be exposed to two calibrants, it is necessary to remove the current sensing elements from contact with the patient""s blood. However, this is not an acceptable procedure in most clinical situations since it can compromise the patient by, for example, increasing the risk of infection.
To address intensity drift in current sensing systems, several referencing schemes have been taught in the art. One commonly practiced approach is to incorporate a fluorescence decay constant that corrects for drift resulting from photo-degradation. This approach is used in correcting for drift in the potassium sensor system taught in Bentsen U.S. Pat. No. 5,958,782 and for correcting drift in the oxygen sensor system taught in Nagel U.S. Pat. No. 5,409,666, both incorporated commercially as part of the LED based sensor system taught in Bentsen U.S. Pat. No. 6,009,339. Each of these patents is incorporated in its entirety herein. Such an approach is insufficient for single fiber sensing elements where photodegradation can be more dramatic (as much as 40% declined in intensity) than in the cassette format and where intensity fluctuations associated with fiber bending are also more pronounced (as much as 60% fluctuations in intensity).
Surgical and clinical environments impose stringent constraints for precision and drift of blood oxygen sensor systems as shown below:
To achieve such precision, the Stern-Volmer quenching constant KSV for an oxygen sensing system is preferably between 0.006 mmxe2x88x921 (Io/Iair=2) and 0.05 mmxe2x88x921 (Io/Iair=9), more preferably between 0.0075 mmxe2x88x921 and 0.02 mmxe2x88x921 (Io/Iair=4.2), and most preferably, between 0.009 mmxe2x88x921 and 0.015 mmxe2x88x921. As discussed by Wolfbeis in Fiber Optic Chemical Sensors and Biosensors, Vol II, CRC Press 1991 and taught by Mauze in U.S. Pat. No. 5,057,277, when using intensity or lifetime measurements to determine analyte concentration, too large a Stern-Volmer quenching constant can be undesirable. In particular, when the quenching constant is too large, relatively large changes in lifetime or intensity values occur over a narrow range of analyte concentrations. At larger analyte concentrations of interest, analyte dependent changes in the fluorescence intensity and lifetime become undesirably small. These considerations are especially problematic in the proper design of a sensor system for monitoring oxygen partial pressure in blood, where accuracy is desired over the range of pO2=40xe2x88x92120 mm Hg, more preferably over the range of 40-180 mm Hg.
Accordingly, for use in the bedside market, there is a need for oxygen sensor systems having calibration plots with slopes or slopes and intercepts that are insensitive to drift and instability caused by variations in fluorescence lifetime, and that can operate within the range required in the clinical environment while satisfying the above specifications for drift and precision for a period of up to 72 hours. There is also a need for oxygen sensor systems that can support rapid (under 5 minute) calibration for pO2. There is also a need for oxygen sensing elements which are capable of being incorporated into a flow-through cassette based sensor system that is compact and light weight. Oxygen sensing elements which avoid leaching out of the fluorescent indicator into the body fluid or tissue are especially desirable.
The present invention provides sensor systems and methods for determining the concentration of oxygen and oxygen-related analytes in a medium, particularly an aqueous-based medium such as blood or body tissue. In one broad aspect, the present sensor systems comprise a sensing element, an excitation assembly, a detector assembly, and a processor assembly, wherein the sensing element comprises a solid polymeric matrix material that is permeable to oxygen or an oxygen related analyte and an indicator that is covalently bonded to the solid polymeric matrix material. The indicator is a luminescent platinum group metal polyaromatic chelate complex capable of having its luminescence quenched by the presence of oxygen. Platinum Group metals are Group VIIIA in the periodic table. The polyaromatic complex comprises three ligands, at least one of which is a bidentate diphenylphenanthroline. The polyaromatic complex is distributed substantially homogenously throughout the matrix material and is covalently bonded to the matrix material via a linker arm. The linker arm is attached to a phenyl group of a diphenylphenanthroline ligand and to the backbone of the polymeric matrix material.
In a particularly useful embodiment, the complex has the formula.
M+L1L2L3
wherein M+ is Ru2+, Os2+, Ir3+, or Rh3+. The ligands L1 and L2 are ide and represent an optionally substituted bidentate phenanthroline or diphenylphenanthroline ligand or an optionally substituted cyclometallated bidentate phenylpyridine ligand or a benzo[h]quinoline ligand. The ligand L3 is a bidentate diphenylphenanthroline ligand substituted by a linker arm which covalently links the metal complex to the matrix material. The linker arm comprises a group selected from the group consisting of a covalent bond, O, C(O)O, an optionally substituted methylene group, an optionally substituted carbon chain comprising 2-20 carbon atoms, and combinations thereof. The carbon chain optionally comprises one or more of the following moieties or combinations thereof: an oxygen atom, a nitrogen atom, a sulfur atom, a silicon atom, a heterocyclic group, and an aryl group. Advantageously, in such a sensing element, the indicator is homogeneously distributed throughout the solid polymeric matrix, a feature that results in linear and reproducible calibration plots.
Any suitable polymeric matrix material may be employed in the sensing element, provided that it functions as described herein. The matrix material, or the precursor thereof, should preferably be such as to chemically react with the linker arm of the indicator and produce a sensing element with a covalently bonded indicator.
Although various polymers can be employed as the matrix material, it is preferred that the matrix material be a silicone-based polymer. Particularly useful polymeric matrix materials include those based on addition cure silicone polymers. If a silicone-based polymer is employed in the matrix material, it may include polymers derived from precursors including vinyl terminated polysiloxanes and polyalkyl(aryl)hydrosiloxanes. Such polyalkyl(aryl)hydrosiloxanes include, but are not limited to, those having the formula 
where each of x and y is independently an integer in the range of 1 to about 500 and R is independently selected from the group consisting of H, alkyl, a substituted alkyl, and a phenyl. Such vinyl terminated polysiloxanes have the formula 
where the sum of m and n is in the range of 100-500 and R is independently selected from the group consisting of alkyl, a substituted alkyl, and a phenyl. Preferably, the silicone-based matrix is free from acids and amines that can leach from the sensing element and change its performance. For sensing oxygen concentrations that are found in blood, it is preferred that a major portion of the R groups are methyl groups. Preferably, the linker arm of the complex is attached to the silicone based polymer by a siloxane or silane linkage.
We have unexpectedly found that the combination of the functionalized indicators having L3 as a bidentate diphenylphenathroline ligand substituted with a linker arm and silicone based matrix materials as described above give rise to oxygen sensing elements wherein the indicator is well dispersed and covalently attached such that the calibration slopes are highly reproducible. Sensor compositions and sensing elements made with these materials overcome several problems associated with ruthenium based oxygen sensing elements of the prior art. In particular, compositions of the extant invention give rise to sensing elements that exhibit long fluorescence lifetimes, in excess of 1 xcexcsec. Furthermore, aggregation of the indicator is minimized, giving rise to substantially linear Stern-Volmer slopes that are consistently greater than 0.009 mmxe2x88x921 and substantially uniform over the range of oxygen partial pressures of 40-180 mm Hg. In addition, the Stern-Volmer slope can be reproducibly controlled through selection of the ratio of methyl to phenyl substituents in the siloxane polymer.
In one preferred embodiment, the solid polymeric matrix material is a dimethylsiloxane polymer or a phenyl-methylsiloxane polymer which is permeable to oxygen and the indicator is a luminescent platinum group metal complex comprising at least one bidentate diphenylphenanthroline ligand having a linker arm that covalently attaches to the polymer backbone. The emission from the complex in this matrix is characterized by a bimolecular quenching rate constant kq for quenching by oxygen and by one or more fluorescence lifetimes xcfx84o above a lowest lifetime xcfx84oL=1 xcexcsec in the absence of oxygen, such that the Stern-Volmer quenching constant KSV is greater than 0.006 mmxe2x88x921 (Io/Iair=2), more preferably greater than 0.0075 mmxe2x88x921, most preferably greater than 0.009 mmxe2x88x921, and substantially uniform over the range of oxygen partial pressures of 40-180 mm Hg.
The excitation assembly of the sensor system is positioned and adapted to provide an excitation signal to the sensing element. The excitation assembly comprises a light source that is preferably selected from the group consisting of light emitting diodes, laser diodes, frequency doubled laser diodes, and solid state light sources. The detector assembly is positioned and adapted to detect the analyte dependent signal from the sensing element and to provide a corresponding electrical signal that can be analyzed by the processor assembly.
A processor assembly is in communication with the detector assembly. The processor assembly includes memory to store information for characterizing a calibration relationship between analyte concentration and a concentration dependent parameter. The processor assembly processes the detected signals to derive the concentration dependent parameter, and provides output signals representative of analyte concentration as a function of the derived concentration dependent parameter and the stored information.
In a number of particularly useful embodiments, referred to hereinafter as xe2x80x9cphase modulation sensorsxe2x80x9d, the sensor system is configured for phase-modulation detection. In phase modulation sensor systems, the signal emitted from the sensing element is intensity modulated, preferably sine wave modulated. This may be done, for example, by exposing the sensing element to an intensity modulated excitation signal or signals. The detector assembly is adapted to sample separately the modulated excitation signal and the modulated signal emitted from the sensing element.
In a first embodiment of the phase modulation sensors, the processor assembly is adapted to determine the extent of the phase shift between the modulated excitation signal and the modulated emission signal. The extent of this phase shift is dependent on the concentration of the analyte in the medium.
In a second embodiment of the phase modulation sensors, the processor assembly is adapted to determine the magnitude of the ratio of demodulation between the modulated excitation signal and the modulated emission signal. The extent of this ratio of demodulation is dependent on the concentration of the analyte in the medium.
In a third embodiment of the phase modulation sensors, the processor assembly is adapted to determine the magnitude of the extent of the phase shift between the modulated excitation signal and the modulated emission signal. The intensity modulated excitation signal is adjusted in frequency so as to maintain a fixed phase shift difference between the modulated excitation signal and the modulated emission signal. The excitation frequency necessary to maintain a fixed phase shift between the modulated excitation and emission signals is dependent on the concentration of the analyte in the medium.
Preferably the excitation assembly, detection assembly, and processor assembly of the phase modulation sensor system are configured to operate at one or more modulation frequencies not to exceed 1 MHz, more preferably not to exceed 500 kHz, and most preferably, not to exceed 200 kHz, such that the sensor system can operate sufficiently within the condition [(kq[O2])2+xcfx892]xcfx84o2 greater than  greater than 1+2kqxcfx84o[O2], where [O2] is the concentration of the oxygen in the sensing element. [O2] can be estimated by the product apO2. This allows the slope of the relationship between the concentration dependent parameter and analyte concentration to be independent of xcfx84o variability for all analyte concentration within the operating range and for all lifetimes xcfx84o greater than xcfx84oL=1 xcexcsec. Preferably, the sensing element and the excitation signal are configured so as to provide an operating condition where the ratio [(kq[O2])2+xcfx892]xcfx84o2/(1+2kqxcfx84o[O2]) exceeds 4, more preferably 6, and most preferably 10. Under these conditions, a constant calibration slope can be achieved and a rapid-single point calibration of the sensor is possible.
A still further broad aspect of the present invention is the provision of a sensor composition and sensing elements useful for sensing the concentration of an oxygen-related analyte in an medium. The present invention also relates to methods of making the sensor composition and methods of using the sensor systems of the present invention to determine blood oxygen levels in patients.
The present sensor systems provide accurate, reliable and reproducible oxygen concentration determinations over the oxygen concentration ranges normally found in human blood. In addition, the present sensor systems provide oxygen concentration determinations that are relatively drift free over a 72 hour period of time. The sensing elements of the present invention further can be manufactured to provide a reproducible calibration slope such that a rapid one-point calibration of the sensing element can be achieved.
These and other aspects and advantages of the present invention are set forth in the following detailed description and claims, particularly when considered in conjunction with the accompanying drawings.